Pore-scale hydrodynamics influence the spatial evolution of bacterial biofilms in a microfluidic porous network

Bacteria occupy heterogeneous environments, attaching and growing within pores in materials, living hosts, and matrices like soil. Systems that permit high-resolution visualization of dynamic bacterial processes within the physical confines of a realistic and tractable porous media environment are rare. Here we use microfluidics to replicate the grain shape and packing density of natural sands in a 2D platform to study the flow-induced spatial evolution of bacterial biofilms underground. We discover that initial bacterial dispersal and grain attachment is influenced by bacterial transport across pore space velocity gradients, a phenomenon otherwise known as rheotaxis. We find that gravity-driven flow conditions activate different bacterial cell-clustering phenotypes depending on the strain’s ability to product extracellular polymeric substances (EPS). A wildtype, biofilm-producing bacteria formed compact, multicellular patches while an EPS-defective mutant displayed a linked-cell phenotype in the presence of flow. These phenotypes subsequently influenced the overall spatial distribution of cells across the porous media network as colonies grew and altered the fluid dynamics of their microenvironment.